1264
Y. Tomabechi et al. / Bioorg. Med. Chem. 18 (2010) 1259–1264
9. Yamamoto, N.; Ohmori, Y.; Sakakibara, T.; Sasaki, K.; Juneja, L. R.; Kajihara, Y.
Angew. Chem., Int. Ed. 2003, 42, 2537.
10. Hojo, H.; Matsumoto, Y.; Nakahara, Y.; Ito, E.; Suzuki, Y.; Suzuki, M.; Suzuki, A.;
Nakahara, Y. J. Am. Chem. Soc. 2005, 127, 13720.
11. Kajihara, Y.; Yoshihara, A.; Hirano, K.; Yamamoto, N. Carbohydr. Res. 2006, 341,
1333.
the mixture and washed with ether. The aqueous phase was ana-
lyzed by MALDI-TOF MS. Transglycosylated B chain 10: MALDI-
TOF MS Calcd for C253H384N47O111S2 [MꢁH]ꢁ 5920.53. Found: m/z
5921.17.
12. Yamamoto, N.; Takayanagi, A.; Yoshino, T.; Kajihara, Y. Chem. Eur. J. 2007, 13,
613.
4.9. Trypsin treated mono-transglycosylated insulin 8
13. Yamamoto, N.; Tanabe, Y.; Okamoto, R.; Dawson, P. E.; Kajihara, Y. J. Am. Chem.
Soc. 2008, 130, 501.
14. Katayama, H.; Utsumi, T.; Ozawa, C.; Nakahara, Y.; Hojo, H.; Nakahara, Y.
Tetrahedron Lett. 2009, 50, 818.
Trypsin digested at the carboxyl side of Arg B22 and Lys B29 in B
chain. A solution of 8 in 2% trypsin containing 0.1 M Tris buffer (pH
7.8, 380 lL) was placed in an Eppendorf centrifuge tube at 37 °C for
15. Sato, M.; Furuike, T.; Sadamoto, R.; Fujitani, N.; Nakahara, T.; Niikura, K.;
Monde, K.; Kondo, H.; Nishimura, S.-I. J. Am. Chem. Soc. 2004, 126, 14013.
16. Yamamoto, K. In Endoglycosidases: Biochemistry, Biotechnology, Application;
Endo, M., Hase, S., Yamamoto, K., Takagaki, K., Eds.; Kodansha: Tokyo, 2006; pp
55–83.
17. Kadowaki, S.; Yamamoto, K.; Fujisaki, M.; Izumi, K.; Tochikura, T.; Yokoyama, T.
Agric. Biol. Chem. 1990, 54, 97.
18. Yamamoto, K.; Kadowaki, S.; Fujisaki, M.; Kumagai, H.; Tochikura, T. Biosci.,
Biotechnol., Biochem. 1994, 58, 72.
3 h. The tube was stopped and heated at 100 °C for 30 min to com-
pletely denature the trypsin. After cooling, the reaction mixture
was analyzed by MALDI-TOF MS. Transglycosylated [A(1–
21) + B(1–22)] 11: MALDI-TOF MS Calcd for C303H468N63O135S6
[MꢁH]ꢁ 7341.00. Found: m/z 7343.35.
19. Haneda, K.; Inazu, T.; Yamamoto, K.; Kumagai, H.; Nakahara, Y.; Kobata, A.
Carbohydr. Res. 1996, 292, 61.
Acknowledgment
20. Yamamoto, K.; Kadowaki, S.; Watanabe, J.; Kumagai, H. Biochem. Biophys. Res.
Commun. 1994, 203, 244.
We would like to thank Professor Kenji Yamamoto from Kyoto
University for his kind discussion. We would also like to thank
the Tokyo Chemical Industry Co., Ltd for providing Endo-M and
SGP. Part of this work was supported by a grant for Hi-Tech Re-
search from Tokai University.
21. Osumi, K.; Makino, Y.; Akaike, E.; Yamanoi, T.; Mizuno, M.; Noguchi, M.; Inazu,
T.; Yamamoto, K.; Fujita, K. Carbohydr. Res., 2004, 339, 2633.
22. Yamamoto, K. In Endoglycosidases: Biochemistry, Biotechnology, Application;
Endo, M., Hase, S., Yamamoto, K., Takagaki, K., Eds.; Kodansha: Tokyo, 2006; pp
129–140.
23. Mizuno, M.; Haneda, K.; Iguchi, R.; Muramoto, I.; Kawakami, T.; Aimoto, S.;
Yamamoto, K.; Inazu, T. J. Am. Chem. Soc. 1999, 121, 284.
24. Haneda, K.; Inazu, T.; Mizuno, M.; Yamamoto, K. Methods Enzymol. 2003, 362,
74.
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